This R21 proposal is aimed at the development of a Drosophila-based bioassay-guided fractionation for the discovery of peptidic natural products that target the CNS. We will screen fractions from the venom of two cone snail species, Conus regius and Conus brunneus, as sample cases for development of the Drosophila-based assay. Components of the venom of cone snails elicit a wide range of neurophysiological responses and show great potential as neuronal probes. The biochemical diversity of the venom is part of a neurochemical strategy used by these animals to compete favorably for prey in the marine environment. The venom of C. regius and C. brunneus are classical examples of this molecular diversity and are particularly suited for discovery, as they are related species, and for one of them (C. regius) we have already produced a significant library of novel conotoxins from biochemical-based fractionations. The main contribution from this project would be to investigate the suitability of Drosophila-based screenings for the discovery of novel conotoxins. This methodology can be applied to the discovery of other supply-limited natural products that can elicit neuromodulatory effects. This is to be done in a multidisciplinary environment for a project that ranges from marine science to Drosophila neurobiology by the collaborative efforts of the Mari and Godenschwege laboratories. These screenings can provide a novel set of neuronal probes or pharmacological agents with unique favorable medicinal properties. As part of the exploratory/developmental phase of our research program and within the R21 program objetives, we seek funding to carry out the development of bioscreening techniques of these venoms and at the same time, we are poised to discover novel conotoxins, assess their biological targets, and evaluate their potential applications as neuronal probes or drug leads. Specifically, we propose to accomplish the following goals: 1) To carry out in vivo functional screening of fractions of the venom of C. brunneus (an Eastern Pacific worm-hunting cone snail species) using electrophysiological measurements on Drosophila melanogaster. The specific neuromodulatory effects observed during the screenings can help to define their specific molecular target type 2) We will use an assortment of bioanalytical techniques to isolate and determine the primary structure of the compounds that show bioactivity during the screenings 3) To compile a Drosophila-based conotoxin library of active components and evaluate their potential as probes for neurobiology applications. Once this library is compiled, structure-activity relationships can be used to established the precise biological role of these compounds. PUBLIC HEALTH RELEVANCE: Peptidic natural products are powerful neuropharmacological agents that can be used for a wide variety of applications. The venom of cone snails, a genus of predatory marine mollusks, are an extremely prolific source of peptidic natural products. Our laboratory (dedicated to study the venom of cone snails) has join forces with the Godenschwege lab to develop new methodologies that will enhance our ability to discover new classes of peptidic natural products. More specifically, we will carry out rapid electrophysiological measurements in tiny Drosophila that consume minimal amounts of precious material isolated from their natural sources. We have chosen the venom of two related cone snail species as a test case for the development of these Drosophila-based methods. We will molecularly identify the bioactive components from these screens and will establish structure-activity relationships that will lead to further studies, such as mammalian-based assays. We are uniquely poised to develop this great potential for "Drug Discovery" as we have already worked-up the biochemical composition of the venom of these snails and immediate correlations between activity and structure can be drawn. Our ultimate goal is to augment the current library of conotoxins with known activity in order to evaluated their potential use as therapeutic agents.